The research described in this proposal focuses on the cellular mechanisms involved in the release of neurotransmitter at a chemical synapse. Since calcium ions are essential to neurotransmitter release, a major theme of the research is the regulation of intracellular calcium by the nerve terminal under various physiological and pathological conditions. Given the central importance of synaptic transmission both to the normal functioning of the nervous system and to pathological disturbances of this function, it is essential that this process be understood in detail. The experiments outlined in this proposal will be performed on the nerve-muscle synapse in the ceratomandibularis muscle of the lizard, Anolis carolinensis. This muscle is very thin and flat, and the presynaptic nerve endings contain large boutons. Using focal extracellular recording, four separate currents -- including a voltage-dependent calcium current -- can be measured in the presynaptic terminal. This allows measurements of transmitter release, made by conventional intracellular recording, to be directly compared to the presynaptic currents responsible for eliciting the release of neurotransmitter. With such information, it will be possible to directly address several issues related to the initiation and regulation of neurotransmitter release from a vertebrate nerve terminal. Specifically, the work described in this proposal is organized around three questions which have been formulated to exploit the inherent optical advantages of the ceratomandibularis muscle and the unique opportunity to monitor changes in calcium current and neurotransmitter release from the same nerve terminal. (1) How do ATP and/or adenosine modulate neurotransmitter release at the vertebrate neuromuscular junction? (2) What is the cellular basis of the defect in neuromuscular transmission that develops in humans with Lambert-Eaton myasthenic syndrome (LEMS)? (3) What is the precise spatial and temporal distribution of calcium in an active presynaptic nerve terminal?